Abstract
Here, we establish a methodology for large-scale quantitative proteomics using SIL (stable isotope labeling) to examine protein expression changes in trophozoite stages of the malaria parasite Plasmodium falciparum following drug treatment. For this purpose, exposure to 13C6 15N1-isoleucine was optimized in order to obtain 99 % atomic enrichment. Proteome fractionation with anion exchange chromatography was used to reduce sample complexity and increase quantitative coverage of protein expression. Tryptic peptides of subfractions were subjected to SCX/RP separation, measured by LC-MS/MS, and quantified using the software tool Census. In drug-treated parasites, we identified a total number of 1,253 proteins, thus increasing the overall number of proteins so far identified in the trophozoite stage by 30 % in the previous literature. A relative quantification was obtained for more than 800 proteins. About 5 % of proteins showed a clear up- or downregulation upon drug treatment.
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Acknowledgement
The authors wish to thank the Alexander von Humboldt Foundation for funding Dr. Judith Helena Prieto. The work was supported by the Deutsche Forschungsgemeinschaft SPP 1710, BE1540/23-1 (to K.B.).
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Prieto, J.H., Fischer, E., Koncarevic, S., Yates, J., Becker, K. (2015). Large-Scale Differential Proteome Analysis in Plasmodium falciparum Under Drug Treatment. In: Peacock, C. (eds) Parasite Genomics Protocols. Methods in Molecular Biology, vol 1201. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1438-8_17
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DOI: https://doi.org/10.1007/978-1-4939-1438-8_17
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